The goal of the research is to characterize the structure and function of the human centromeric region. The centromere is the cytologically visible and genetically defined region of human chromosomes which is responsible for proper segregation of chromosomes in both mitosis and meiosis. These processes involve specialized chromosomal DNA sequences which interact specifically with cellular components. To investigate those DNA sequences that are required for centromeric activity the following research plan is proposed: 1. Sequences from the regions around the centromere of specific human chromosomes will be isolated by screening chromosome sorted libraries with 308, a human DNA sequence from chromosome 6, but homologous to centromeres of all human chromosomes. DNA from centromeric regions flanking the 308 related sequences will be obtained from screening recombinants for overlapping sequences. In this way unique as well as repetitive sequences will be obtained. 2. Centromeric sequences will be characterized to determine chromosomal localization by in situ hybridization to metaphase chromosomes and organization by Southern blot analysis and nucleotide sequence analysis. 3. To test for centromeric function, human DNA sequences will be inserted into viral vectors which usually integrate into chromosomes and transform mammalian cells. In these cells, sequences which function as centromeres will permit vectors to be stably maintained and segregate correctly with each cell division. The extrachromosomal localization will be determined by in situ hybridization and the organization of these sequences will be analyzed by Southern blot analysis. 4. Robertsonian translocation chromosomes will be analyzed with respect to the organization of their centromeric regions by using in situ hybridization, Southern blot analysis, and nucleotide sequence analysis. 5. Centromeric DNA probes will be used as cytological markers in in situ hybridization studies to determine the arrangement of chromosomes in the interphase nucleus.